In this project the basic structures of the proteins that are responsible for biological force generation are being studied. The contractile proteins from skeletal and cardiac muscle are studied first and become a frame of reference for comparative studies on related proteins from other tissues including smooth muscle, brain, blood platelets, and Acanthamoeba. Work directed toward determination of the complete amino acid sequence of the myosin heavy chain is concentrated upon large fragments obtained by chemical and limited enzymic cleavage of myosin. One of these, a 22,000 dalton peptide, is the COOH terminus of myosin, and is entirely alpha-helical; determination of its sequence may give information regarding the mechanism of assembly of the thick filament of muscle. Another fragment under investigation, a 51,000 dalton tryptic fragment, contains one residue of e-N-trimethyllysine, and, together with a 10,000 dalton peptide sequenced earlier, constitutes most of the enzymically active head portion of myosin. Comparative studies will be focused upon 1) 10,000 dalton peptide from cardiac myosin; this fragment contains residues that are probably part of the catalytic site, and we will attempt to compare the normal cardiac peptide with the equivalent fragment from hearts of thyrotoxin animals, and 2) the 22,000 dalton helical peptide from smooth muscle myosin, since it aggregates differently, and the molecular basis for the difference may be recognizable from the comparison. Studies of actins include completion of the sequence of Acanthamoeba actin and identification of sidechains that are involved in the binding of ADP to actin. The latter studies involve covalent crosslinking of nucleotide and protein by intense illumination. Sequence studies on the light chains of human cardiac myosin will be continued toward the goal of comparing the normal chains with equivalent protein from other myosins, and establishing whether in hypertrophic hearts there exist genetic or posttranslational alterations in structure.